Project 3: The diagnostic methods currently used in clinical microbiology have limited capabilities to identify pathogens in a rapid turn-around time that allows prompt initiation of a therapeutic intervention and appropriate infection control measures. This is especially relevant when facing novel emerging and reemerging pathogens and potential biothreat agents. Rather than focusing on the identification of hundreds of potentially pathogenic agents in a traditional microbe-based approach, we propose a comprehensive analysis of the host response to different classes of pathogens as a novel diagnostic tool. Based on our preliminary studies in individuals with acute infections, we hypothesize that each class/group of microorganisms induces a host response pattern [in the immune system] which defines a characteristic biosignature that can be used for diagnostic purposes. Using microarray gene expression analysis we will determine: a) whether there are infection-induced specific biosignatures which are different than those induced by other non-infectious conditions and/or stressors such as autoimmune diseases, cancer, trauma, and transplantation; b) whether this biosignature can identify different classes of pathogenic microorganisms, and c) whether the biosignature can be used as a prognostic indicator of disease severity in different types of infections. Aim 1 will determine whether Gram positive and GRam negative bacteria yield different biosignatures in human blood in vivo. Aim 2 will determine whether gene expression in human blood will permit to distinguish between bacterial and viral infections. Aim 3 will determine whether established biosignatures permit diagnosis of "unknown infection". Aim 4 will determine which immune cells in blood carry the microbe biosignature. This study will establish the diagnostic and prognostic value of pathogen biosignatures in vivo. Our goal is to establish microarray analysis of blood leukocytes as a platform technology for the diagnosis of biothreat exposure and decision making with regard to therapeutics and, on a larger scale, Public Health.